Testicular teratomas arise in humans as benign, non-seminomatous germ cell tumors characterized by the differentiation of a diverse array of specific cell and tissue types. These unusual tumors reflect the close relationship between germ cells, embryonic stem cells, and tumor cells, and hold important clues about the mechanisms that program these cell fates and regulate the transitions between them. Although teratomas were virtually unknown in rodent models, a spontaneous mouse mutation, Ter, was discovered in the 1970s that led to a high incidence of testicular teratomas, specifically on the 129/SvJ genetic background but not on other strains such as C57BL/6J. On most genetic backgrounds Ter-/- XY germ cells disappear completely by the time of birth. Recently, the Ter mutation was mapped to Dnd1, the mouse ortholog of a gene involved in germ cell regulation in zebrafish. DND1 is an RNA-binding protein that targets several known transcripts involved in germ cell differentiation and cell cycle control and protects them from miRNA-mediated degradation. Based on our preliminary data, we hypothesize that DND1 regulates many targets that control germ cell programming at transitional stages of PGC development. We propose to use an unbiased Y3H screen to identify the mRNA clients of DND1, and a computational approach to identify interacting miRNAs and possible strain differences that may account for the different sensitivity to teratoma formation. We also plan to create a conditional tagged allele of Dnd1 and delete it (1) at germ cell specification/migration stages, (2) during the fetal stage of germ cell transition to pro- spermatogonia in the testis and (3) in postnatal animals. We will use this tagged allele to determine the cellular localization of DND1 during development, and to immunoprecipitate DND1 bound to its in vivo mRNA targets. We will also isolate wild type and mutant germ cells at E14.5 from both the susceptible (129) and non-susceptible (B6) strains to obtain global expression profiles. These data will be correlated with DND1 target data to elucidate the transcriptional and post-transcriptional mechanisms governing germ cell cycle progression and strain sensitivity to tumor formation. The integration of translational mechanisms with transcriptional networks underlying strain sensitivity to tumor formation has immediate clinical relevance to human germ cell tumors.
Germ cells are crucial to reproductive health. Disruptions of their development lead to sterility or the formation of germ cell tumors. By integrating levels of transcriptional and post-transcriptional information, we will identify the mechanisms through which DND1, an RNA-binding protein, controls the cell cycle and programming transitions of germ cells. Although DND1 is specific to germ cells, we anticipate that the regulatory mechanisms involved may be common to many pluripotent stem cell populations.
